Heterogeneous binding assays require that a means be provided to separate a labeled binding reagent from an unlabeled binding reagent. Frequently, a surface is provided to which is bound a specific binding ligand or receptor. Various surfaces have been used, such as latex beads, which can be filtered; tubes or wells, usually plastic, which also serve as the container for the assay mixture; magnetic particles which can be separated in a magnetic field gradient; insoluble polymers which are separated by centrifugation or are used as the stationary phase of a chromatograph; bibulous materials such as cellulose or glass paper through which reagents can be filtered or transferred by capillary action.
U.S. Pat. No. 4,659,678 describes a method for carrying out an immunoassay in which the complex is formed in a liquid medium prior to binding to a solid support by the use of one or more monoclonal antibodies. Detection of the bound sample is measured using labeling methods such as labeling with radioactive iodine, and fluorimetric and enzyme labeling.
U.S. Pat. No. 4,780,423 describes a heterogeneous assay using controlled pore glass particles. The controlled pore glass particles are used in a fluorescent immunoassay as the support for the specific binding partner bound to a ligand. As used in the invention, the glass particles bind a complex of interest, the detection of which is achieved by use of a fluorescent probe. Measurement of fluorescence is carried out in the presence of the glass particles.
U.S. Pat. No. 4,298,685 describes the use of biotin-labeled antibodies for the quantitative determination of a biological substance in a test sample. Quantitative measurements of the amount of biological sample present are obtained by the use of an enzyme label, such as horseradish peroxidase, which when bound to the sample can be used as a means to detect the presence of the biological substance in the sample.
The use of solid particles, such as magnetic particles or glass beads, to serve as the support for an immunologic assay is known. An example of such assays include the use of magnetic particles as the solid support in a fluorometric immunoassay as described in U.S. Pat. No. 4,777,145. The use of avidin-coated glass beads in immunoaffinity chromatography and a method for preparing such avidin-coated beads is described by Babashak J. V. and T. M. Phillips, J. of Chromatography 444:21 (1988).
The present invention provides a means to carry out the various heterogeneous binding assays using the improved method of the invention so as to achieve high capacity, rapid binding and convenient washing of the stationary phase of the heterogeneous assay without centrifugation or conventional filtration. In general, conventional filtration systems require expensive membranes which are inconvenient and often cannot be reused. Centrifugation is inconvenient to automate, and tubes or wells do not offer adequate surface area or geometry to provide a high binding capacity and rapid binding.